Fig. 3
SARS-CoV-2 infection of human PCLS is leading to a low level of replication. A ACE2 and TMPRSS2 mRNA levels per 18S rRNA in PCLS of selected donors. Boxplots indicate the median value (centerline) and interquartile ranges (box edges), with whiskers extending to the lowest and the highest values. (ACE2, n = 15; TMPRSS2, n = 4). Fold difference was calculated by comparing the median ACE2 expression of all the PCLS (n = 15) and WD-NEC (n = 3) donors tested. B PCLS were infected one to seven days after generation with 106 PFU of SARS-CoV-2 (SARS-CoV-2/München1.1/2020/929; Early isolate) and Delta isolate (Delta AY.127, hCoV-19/Switzerland/BE-IFIK-918–4879/2021). After 2 h PCLS were washed twice with PBS. Growth medium was changed daily. 4, 24, 48, and 72 h p.i. supernatants and PCLS were harvested for further analysis. C Histological section of a representative Mock- and SARS-CoV-2 Delta isolate-infected PCLS 48 h p.i., stained for the nucleocapsid (N) protein and counterstained for hematoxylin. D Representative confocal microscopy evaluation of Mock- and SARS-CoV-2 Early isolate-infected PCLS 48 h p.i., stained for DAPI (grey) and SARS-CoV-2 N protein (green). a, Mock; b-e, SARS-CoV-2 Early isolate-infected PCLS. a-c, micrographs are projections of 3D captures. a-e, magnification 10X. E Supernatants of infected PCLS were analyzed with a PFU assay. Early isolate, n = 18; Delta isolate, n = 7. F Extracellular viral RNA in the supernatant of SARS-CoV-2 Early isolate-infected PCLS over time expressed as viral RNA copies per ml (n = 14). G Tissue-associated viral RNA loads per 18S rRNA in PCLS over time following SARS-CoV-2 infection. Dotted line represents the detection limit. Early isolate, n = 15; Delta isolate, n = 7 H Correlation of tissue-associated viral RNA and donor age. I Correlation of tissue-associated viral RNA and deltaCt CDKN2A (p16) (deltaCt = Ct CDKN2A-Ct B2M). Association was tested using the Spearman rank correlation test. Each symbol represents one individual donor at 24 h (circle), 48 h (square), or 72 h p.i. (triangle)